The inner retina is a significant source of lactic acid, consistent with the high lactate concentration (3.8 - 13 mM) at the SRS even in light-adapted eyes. Lactic acid is a byproduct of anaerobic respiration, and is released in high quantities in the dark adapted eye. This increase in lactate release in the dark is mainly caused by (1) increased glucose metabolism at the outer retina;(2) reduced retinal oxygen level, which triggers anaerobic respiration;(3) glutamate-induced lactate release from Muller cells. Accumulation of lactic acid within the subretinal space is detrimental to the function of the photoreceptors. Mice lacking lactic acid removal mechanisms gradually lose their vision weeks after birth. The Retinal Pigment Epithelium (RPE) is strategically located between the photoreceptor outer segments and the choroidal blood supply. The RPE supports photoreceptor function in multiple ways, one of which is metabolite (CO2/HCO3, and lactic acid) removal from the subretinal space. To remove lactic acid from the subretinal space, the RPE expresses lactate transporters, of the MCT family of monocarboxylate transporters, at the apical (MCT1) and basolateral membranes (MCT3 and MCT4). This study uses pHi-imaging techniques, combined with the more traditional electrophysiological methods of epithelial voltage and resistance measurements to study the mechanisms involved in lactate transport in the RPE. Earlier, our experiments show that perfusing Ringer solution containing 20 mM lactate to the apical bath caused a pHi response with two distinct phases: (1) a fast acidification (phase 1), followed by (2) a slow alkalinization (phase 2). Subsequent experiments demonstrate that the fast acidification (phase 1) was mediated by H/Lac entry into the cell via MCT1 at the apical membrane, whereas the slow alkalinization (phase 2) was caused by the acidification induced activation of the Na/H exchanger. Our earlier data also shows that apical lactate also caused an increase in TEP and an increase in total tissue resistance (Rt). A change in TEP is indicative of a change in membrane voltage, mediated by an activation or inhibition of an electrogenic process. Since MCT transporters are electroneutral transporters, this observation suggests that other transport mechanisms are stimulated or inhibited upon apical lactate entry. Earlier, we showed that this TEP response can be partially inhibited by basal application of DIDS, a potent but non-specific Cl-channel inhibitor. This data suggests that Cl-channels at the basolateral membrane are activated upon apical lactate entry. Cl-efflux at the basolateral membrane is known to be mediated mainly by the cystic fibrosis transmembrane conductance regulator (CFTR) and Ca2+ activated Cl channels. However, current experiments suggest that the apical lactate induced TEP response was not caused by the activation of either of these two channels. Therefore we test the possibility that apical lactate may have activated ClC-2, a zinc-sensitive Cl-channel. ClC-2 is much less characterized in the RPE, and its localization and expression within the RPE is unknown. In current experiments, we show that ClC-2 proteins are highly expressed in RPE. In addition, microarray analysis also showed high mRNA expression for the ClC-2 protein. More importantly, basal application of zinc reduced the apical lactate induced TEP response by 30-50%. In contrast, apical application of zinc to the apical surface did not reduce the apical lactate induced TEP response. Collectively, our data suggests that ClC-2 is expressed at the basolateral membrane and mediates, in part, the apical lactate induced TEP response. If the basal application of either zinc or DIDS were unable to completely block the apical lactate induced TEP response, it is possible that part of this TEP response was mediated by a separate process. In support of this notion, the complete removal of Cl in the solution did not completely block the apical lactate induced TEP response. Present experiments show that apical barium was able to inhibit the lactate induced TEP response by 50%, suggesting that apical lactate entry caused potassium efflux across the apical membrane via barium-sensitive K-channels. Further, we show that in the presence of apical barium and basal zinc, the apical lactate induced TEP response was essentially completely blocked, thus confirming that apical lactate entry into the cell stimulated K-channel at the apical membrane and ClC-2 at the basolateral membrane. As a next step, we study the mechanism of activation of these two channels (i.e., K- and Cl- channels) by apical lactate. A possibility is that apical lactate causes the cell to swell, thus stimulating KCl efflux from the cell. To test this notion, we used calcein imaging technique to monitor cell volume changes. These experiments show that apical lactate caused a small increase in cell volume. However, from these experiments, we also eliminated the possibility that this small volume change could stimulate the K- and Cl- channels because sucrose-induced cell swelling did not significantly affect TEP. In another set of experiments, we also show that these K- and Cl- channels were not directly activated by the apical lactate induced acidification. These experiments suggest that the lactate induced activation of K- and Cl- channels may be mediated by allosteric interactions with monocarboxylates. Preliminary experiments have also showed that apical lactate caused a decrease in intracellular calcium concentration. This may have other effects on cell physiology that will be further investigated.